Neuropeptide Y (NPY) is a 36 amino acid peptide that is widely distributed in both the central and peripheral nervous systems (1). As might be expected from its widespread distribution, NPY has a plethora of physiological actions, including effects on blood pressure, hormone release, gut motility, smooth muscle tone, sleep and circadian rhythms, feeding, thermogenesis, neuronal excitability, nociception, cognition, mood and emotional responses. NPY mediates these physiological effects via interaction with at least six distinct G protein-coupled receptors (designated Y.sub.1 -Y.sub.6) (2). It is possible that additional NPY receptors remain to be cloned and characterized. Available data indicate that the NPY Y.sub.5 receptor mediates the effects of NPY on feeding, thermogenesis, neuronal excitability and seizure activity, diuresis, natriuresis and calciuresis (3,4,5).
Several lines of evidence suggest that NPY plays a key role in the control of body weight. Central administration of NPY increases food intake and decreases thermogenesis in satiated animals while reductions in endogenous NPY via antisense oligonucleotide or immunoneutralization techniques leads to a decrease in food intake (1,6,7). As would be expected for an orexigenic peptide, hypothalamic NPY peptide and mRNA levels are increased after fasting and in genetically obese mice (8). In fact, recent experiments with transgenic mice lacking NPY indicate that NPY is required for the maintenance of the obese phenotype of ob/ob mice (9). Conversely, the satiety signal leptin appears to decrease food intake and body weight in part by decreasing NPY synthesis and release (10). The pharmacological properties of the Y.sub.5 receptor subtype most closely match the pharmacological properties of the receptor mediating the effects of NPY on feeding (3). These data suggest that NPY is a key modulator of body weight and that NPY Y.sub.5 receptor antagonists will be useful anti-obesity agents.
NPY has also been shown to block kainic acid-induced seizures in rats, suggesting that NPY may be a potent anti-epileptic agent (4). The pharmacological properties of the receptor that mediate the anti-epileptic effects of NPY are similar to the pharmacological properties of the Y.sub.5 receptor (4). Therefore, NPY Y.sub.5 receptor agonists may be useful anti-epileptic agents.
NPY also elicits a diuretic, natriuretic and calciuretic effect in the kidney (5). Again, the pharmacological properties of the receptor that mediates these effects are similar to the pharmacological properties of the Y.sub.5 receptor (5). It is conceivable, therefore, that NPY Y.sub.5 receptor agonists would be effective anti-hypertensive agents and/or useful in the treatment of disorders of calcium metabolism.
NPY has numerous physiological effects which cannot yet be conclusively ascribed to a particular NPY receptor subtype (1). It is certainly possible that some of these effects are due to interaction of NPY with the Y.sub.5 receptor. The involvement of NPY in sleep and other circadian rhythms suggests that Y.sub.5 receptor agonists might be useful in the treatment of sleep disorders, including jet lag. The known effects of NPY on hormone release suggest that Y.sub.5 agonists or antagonists might be useful as contraceptives or in the treatment of infertility or sexual dysfunction. NPY also regulates vascular tone in cerebral vessels, suggesting that Y.sub.5 agonists or antagonists might have value in the treatment of migraine. Since NPY is also known to be involved in the transmission of painful stimuli, Y.sub.5 agonists or antagonists might be useful analgesics. NPY has also been shown to have effects on cognitive processes and, hence, Y.sub.5 receptor agonists or antagonists might be effective cognition-enhancing agents. Disorders of gut motility could also be treated with a Y.sub.5 agonist or antagonist. NPY itself is also anxiolytic and, thus, Y.sub.5 receptor agonists might be useful anxiolytic agents. Finally, NPY is localized in brain regions that play a role in affective disorders, suggesting that Y.sub.5 ligands could be useful antidepressants or antipsychotic agents.
The present inventors have isolated cDNAs encoding the rat and human NPY Y.sub.5 receptors and used those nucleic acids to construct a novel chimeric cDNA construct that enables high levels of human NPY Y.sub.5 receptor expression.
U.S. Pat. No. 5,603,024 ('024 patent) refers to isolated cDNAs encoding human and rat NPY Y.sub.5 receptors. The cDNA sequences disclosed in the '024 patent, however, differ from the sequences of the cDNAs isolated by the present inventors and used in the construction of the novel chimeric receptor cDNA.
For example, the sequences of the human NPY Y.sub.5 receptor isolated by the present inventors and that disclosed in the '024 patent diverge prior to nucleotide number 82 of the '024 sequence. The net result of this divergence is that the translated human NPY Y.sub.5 receptor protein encoded by the '024 patent cDNA has 10 additional amino acids at the amino terminus relative to the translated protein encoded by the human cDNA isolated by the present inventors. Sequencing of the human NPY Y.sub.5 receptor gene reveals that the sequence disclosed in the '024 patent is in reality a genomic sequence.
Likewise, sequence analysis indicates that the rat NPY Y.sub.5 receptor cDNA sequence disclosed in the '024 patent is in fact a genomic sequence.
Similar conclusions were drawn in the recently published International Application WO 97/17440.
In view of the important role of the NPY Y.sub.5 receptor in many physiological processes and medical conditions, there is a need for materials and methods for identifying selective agonists and antagonists of the NPY Y.sub.5 receptor.